1. Field of the Invention
This invention relates to a composite separating agent, which has a high mechanical strength and an excellent permeability of liquids when packed in a column and thus is suitable for chromatographically separating biopolymers such as proteins, as well as a process for producing the same.
2. Prior Art
Separation and purification of biopolymers such as proteins is highly limited by the nature of the materials to be separated, being different from the case of common low molecular weight materials. That is to say, proteins should be separated under mild conditions since they are liable to be denatured or decomposed when exposed to a high temperature, organic solvents, acids or alkalis.
Accordingly, known agents for separating and purifying biopolymers such as proteins fall within the following types:
(1) a separating agent obtained by crosslinking polysaccharides such as agarose or dextran;
(2) a separating agent obtained by crosslinking polyvinyl alcohol with epichlorohydrin;
(3) a separating agent obtained by copolymerizing an acrylate monomer with a crosslinking polyalkylene glycol polyacrylate monomer;
(4) a separating agent obtained by coating the surface of a separating agent (3) with a hydrophilic material; and
(5) a separating agent obtained by charging an agarose gel within pores of a porous inorganic kieselguhr carrier.
The above separating agents (1) and (2) are particularly effective in gel filtration chromatography since they are highly hydrophilic, hardly cause irreversible adsorption of proteins and have a giant network structure. However these separating agents are non-rigid and thus inferior in mechanical strength, which brings about a serious disadvantage. That is to say, when one of these separating agents is applied to column separation on an industrial scale to be carried out at a high column height and at a high flow rate, the separating agent would be compacted and thus the liquid permeation becomes impossible.
On the other hand, the separating agent (3) is rigid and thus excellent in mechanical strength. However it involves some hydrophobic portion which would often cause irreversible adsorption of proteins. Further the porous structure thereof brings about a low separation performance, compared with those of the separating agents (1) and (2).
The separating agent (3), wherein the surface of a separating agent is coated with a hydrophilic material, hardly causes irreversible adsorption of proteins and is rigid. Thus it is excellent in mechanical strength. However the porous structure thereof results in a low separation performance, compared with those of the separating agents (1) and (2).
The above separating agent (5), wherein an agarose gel is charged within pores of a porous inorganic kieselguhr carrier, is rigid and has a high mechanical strength. However inorganic porous materials are scarcely resistant against acids and alkalis in general. Thus a separating agent comprising a porous inorganic material is highly restricted at the washing step in the separation. Further it has a poor durability.